Page 1. Outline. Photoacoustic Tomography: Multiscale Imaging from Organelles to Patients by Ultrasonically Beating the Optical Diffusion Limit

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1 3 OILAB.SEAS.WUSTL.EDU 2 OILAB.SEAS.WUSTL.EDU OILAB.SEAS.WUSTL.EDU Photoacoustic Tomography: Multiscale Imaging from Organelles to Patients by Ultrasonically Beating the Optical Diffusion Limit Lihong V. Wang Gene K. Beare Distinguished Professor Optical Imaging Laboratory Departments of Biomedical Engineering and Radiology Washington University in St. Louis Photoacoustics@gmail.com Disclosure Statement Lihong Wang Dr. Lihong Wang has disclosed the following financial relationships. Any real or apparent conflicts of interest related to the content of this presentation have been resolved. Affiliation/Financial Interest Consultant/Share Organization Microphotoacoustics, Inc. Outline Motivations and challenges Photoacoustic tomography Photoacoustic computed tomography Photoacoustic microscopy Time-reversal wavefront engineering Compressed ultrafast photography Page

2 Human Organ 6 OILAB.SEAS.WUSTL.EDU Depth Skin Cell 5 OILAB.SEAS.WUSTL.EDU 4 OILAB.SEAS.WUSTL.EDU Motivations for Imaging with Light Light-matter interaction uniquely positioned at the molecular level Electromagnetic spectrum Wavelength: mm µm nm pm Radiowave/microwave Optical X-ray Gamma Non-ionizing (safe) Ionizing (DNA damage) Medical x-rays PET Motivations for Imaging with Light Light-matter interaction uniquely positioned at the molecular level Fundamental role of molecules in biology and medicine In vivo functional imaging analogous to functional MRI In vivo metabolic imaging analogous to PET In vivo molecular imaging of gene expressions or disease markers In vivo label-free histologic imaging of cancer without excision Oxy- & deoxyhemoglobins Brain activation Glucose uptake Melanoma hallmark Photoacoustic microscopy of cell nuclei Source: Wikipedia Challenges in Optical Penetration Photon propagation 0 µm 00 µm mm 2 mm Classical planar optical microscopy Aberration limit: /scattering coefficient Confocal or two-photon microscopy Optical coherence tomography Diffusion limit: 0/scattering coefficient cm Photoacoustic tomography 0 cm m Dissipation limit: 0/attenuation coefficient Wavefront engineering with internal guide stars? Absorption limit: 0/absorption coefficient LV Wang, HI Wu, Biomedical Optics (Wiley, 2007); LV Wang, JJ Yao, Nature Methods 3, 627, 206 Page 2

3 9 OILAB.SEAS.WUSTL.EDU 8 OILAB.SEAS.WUSTL.EDU 7 OILAB.SEAS.WUSTL.EDU Outline Motivations and challenges Photoacoustic tomography Photoacoustic computed tomography Photoacoustic microscopy Time-reversal wavefront engineering Compressed ultrafast photography Photoacoustic Computed Tomography: Deep Penetration with Optical Contrast and Ultrasonic Resolution () ns laser pulse (within safety limit) (2) Absorption of photons (3) Rapid heating (~ mk) (4) Ultrasonic emission: (5) Ultrasonic detection mk 8 mbar (800 Pa), detectable of unscattered phonons (acoustic scattering ~ optical scattering/000) X Wang, Y Pang, G Ku, G Stoica, LV Wang, Nature Biotech 2, 803, 2003 First Functional (Also First In Vivo) Photoacoustic Tomography in Small Animals with Intact Scalp and Skull Left-whisker stimulation Right-whisker stimulation Contralateral hemodynamic response 5 mm Min Max Differential absorption X Wang, Y Pang, G Ku, G Stoica, LV Wang, Nature Biotech 2, 803, 2003 Page 3

4 2 OILAB.SEAS.WUSTL.EDU OILAB.SEAS.WUSTL.EDU Imaging depth (mm) 0 OILAB.SEAS.WUSTL.EDU Number of papers Growth of Photoacoustic Tomography Largest conference since 200 in 20,000-attendee Photonics West Conference Web of Science Number of papers Omniscale In Vivo Photoacoustic (PA) Tomography with Consistent Contrast 0 2 Lateral resolution Axial resolution Depth-resolution ratio = 200 Low-freq PA tomography PA macroscopy Acoustic-resolution PA microscopy (PAM) Optical-resolution PAM Submicron PAM Sub-wavelength PAM PA nanoscopy 0-2 Protein Organelle Cell Tissue Organ Spatial resolution ( m) Omniscale biological research from organelles to small-animal organisms Translation of microscopic lab discoveries to macroscopic clinical practice LV Wang, S Hu, Science 335, 458, 202; LV Wang, Nature Photon 3, 503, 2009 Single Impulse Panoramic Photoacoustic Computed Tomography Hollow cone Lasers Ultrasonic focus Solid cone Brain imaging Diffuser Hollow cone Full-ring ultrasonic array Lei Li, Liren Zhu, Cheng Ma, Konstantin Maslov, LV Wang, unpublished Conical lens Computer : DAQ Trunk imaging Page 4

5 5 OILAB.SEAS.WUSTL.EDU 4 OILAB.SEAS.WUSTL.EDU 6 mm 3 OILAB.SEAS.WUSTL.EDU 50 Hz Frame-Rate Photoacoustic CT of Mice In Vivo: Liver/Portal Vein Region Lei Li, Liren Zhu, Cheng Ma, Konstantin Maslov, LV Wang, unpublished Photoacoustic Computed Tomography of the Whole Brain of a Mouse In Vivo mm Pengfei Zhang,, LV Wang, unpublished Reversibly Switchable Photoacoustic Genetic Imaging of Mouse Brain Tumor ON = signal + BG OFF = BG ON OFF = signal Overlay 0 Photoacoustic amplitude 2 mm [JJ Yao, AA Kaberniuk], L Li, DM Shcherbakova, R Zhang, LD Wang, G Li, VV Verkhusha@Einstein, LV Wang, Nature Methods 3, 67, Norm. BphP concentration Page 5

6 Imaging depth (mm) OILAB.SEAS.WUSTL.EDU 7 OILAB.SEAS.WUSTL.EDU 8 OILAB.SEAS.WUSTL.EDU In Vivo Human Breast Panoramic Photoacoustic Computed Tomography Laser Li Lin, Junhui Shi, Konstantin Maslov,... LV Wang, unpublished Photoacoustic Computed Tomography of Human Breast In Vivo: Volunteer # To chest wall: 3 cm 2 cm 0 cm Left breast cm Right breast Li Lin, Junhui Shi, Konstantin Maslov,... LV Wang, unpublished Omniscale In Vivo Photoacoustic (PA) Tomography with Consistent Contrast 0 2 Lateral resolution Axial resolution Depth-resolution ratio = 200 Low-freq PA tomography PA macroscopy Acoustic-resolution PAM Optical-resolution PAM Submicron PAM Sub-wavelength PAM PA nanoscopy 0-2 Protein Organelle Cell Tissue Organ Spatial resolution ( m) LV Wang, S Hu, Science 335, 458, 202; LV Wang, Nature Photon 3, 503, 2009 Page 6

7 2 OILAB.SEAS.WUSTL.EDU Imaging depth (mm) 20 OILAB.SEAS.WUSTL.EDU 9 OILAB.SEAS.WUSTL.EDU Outline Motivations and challenges Photoacoustic tomography Photoacoustic computed tomography Photoacoustic microscopy Time-reversal wavefront engineering Compressed ultrafast photography First 3D Photoacoustic Microscope z y x Motor driver Tunable laser Photodiode Nd:YAG pump laser Translation stages Amplifier Optical illumination Ultrasonic transducer Conical lens Sample holder AD Mirror Base Computer Heater & temperature controller Dual foci Tissue Donut-shaped illumination K Maslov, G Stoica, LV Wang, Optics Lett 30, 625, 2005 H Zhang, K Maslov, G Stoica, LV Wang, Nature Biotech 24, 848, 2006; Nature Protoc 2, 797, 2007 Omniscale In Vivo Photoacoustic (PA) Tomography with Consistent Contrast 0 2 Lateral resolution Axial resolution Depth-resolution ratio = 200 Low-freq PA tomography PA macroscopy Acoustic-resolution PAM Optical-resolution PAM Submicron PAM Sub-wavelength PAM PA nanoscopy 0-2 Protein Organelle Cell Tissue Organ Spatial resolution ( m) LV Wang, S Hu, Science 335, 458, 202; LV Wang, Nature Photon 3, 503, 2009 Page 7

8 24 OILAB.SEAS.WUSTL.EDU 0 Oxygen saturation 0 20% Photoacoustic signal change 23 OILAB.SEAS.WUSTL.EDU Imaging depth (mm) 22 OILAB.SEAS.WUSTL.EDU In Vivo Photoacoustic Microscopy of the Human Skin En face PAM B-scan 584 nm Epidermal-dermal junction Stratum corneum Epidermis Photo Dermis Subpapillary plexus mm mm C Favazza, O Jassim, LA Cornelius, LV Wang, J Biomed Optics 6, 0605, 20; Collaboration: LA Cornelius Omniscale In Vivo Photoacoustic (PA) Tomography with Consistent Contrast 0 2 Lateral resolution Axial resolution Depth-resolution ratio = 200 Low-freq PA tomography PA macroscopy Acoustic-resolution PAM Optical-resolution PAM Submicron PAM Sub-wavelength PAM PA nanoscopy 0-2 Protein Organelle Cell Tissue Organ Spatial resolution ( m) LV Wang, S Hu, Science 335, 458, 202; LV Wang, Nature Photon 3, 503, 2009 In Vivo Photoacoustic Microscopy of Cerebral Hemodynamic Response to Electric Hindpaw Stimulation 3D imaging rate: Hz mm 500 μm JJ Yao, LD Wang, JM Yang, KI Maslov, TTW Wong, Lei Li, CH Huang, J Zou@TAMU, LV Wang, Nature Methods 2, 407, 205; Featured by Science, doi:0.26/science.aab0393 Page 8

9 27 OILAB.SEAS.WUSTL.EDU 26 OILAB.SEAS.WUSTL.EDU 25 OILAB.SEAS.WUSTL.EDU Oxygen saturation of hemoglobin (so 2 ) Hz B-scan 20 Hz B-scan In Vivo Single-Cell Photoacoustic Flowoxigraphy: Flow and Oxygenation Imaging Human cuticle capillary Single red blood cells (200 Hz B-scan, 20 3D images/s) 20 μm 0.7 LD Wang, K Maslov, LV Wang, PNAS 0, 5759, 203; HC Hsu, LD Wang, LV Wang, JBO 2, , 206 In Vivo Photoacoustic Microscopy and Short-Pulsed Laser Therapy of Single Circulating Tumor Cells Goals:. Remove primary melanoma 2. Clear circulating tumor cells 3. Uncage antigens alive 4. Elicit immunoresponse 5. Destroy metastases Yun He, LD Wang, J Shi, J TAMU, LV Wang, unpublished Label-Free Photoacoustic Histology by Imaging DNA & RNA in Cell Nuclei Photoacoustic microscopy without staining Histology with hematoxylin and eosin staining 20 µm D Yao, R Chen, K Maslov, Q Zhou, LV Wang, J Biomed Optics 7, , 202; Collaboration: Q USC Page 9

10 30 OILAB.SEAS.WUSTL.EDU 29 OILAB.SEAS.WUSTL.EDU Imaging depth (mm) 28 OILAB.SEAS.WUSTL.EDU Label-Free Photoacoustic (PA) Nanoscopy of a Mitochondrion with Sub-Organelle Resolution: Beat Optical Diffraction Nonlinearly PA microscopy (Resolution: 234 nm) PA nanoscopy (Resolution: 90 nm) Comparison Electron microscopy 500 nm A Danielli, K Maslov, A Garcia-Uribe, A Winkler, CY Li, LD Wang, Y Chen, G Dorn, LV Wang, J Biomed Optics 9, , 204; Collaboration: G Dorn; J Yao, LD Wang, CY Li, C Zhang, LV Wang, Phys Rev Lett 2, 04302, 204 Omniscale In Vivo Photoacoustic (PA) Tomography with Consistent Contrast 0 2 Lateral resolution Axial resolution Depth-resolution ratio = 200 Low-freq PA tomography PA macroscopy Acoustic-resolution PAM Optical-resolution PAM Submicron PAM Sub-wavelength PAM PA nanoscopy 0-2 Protein Organelle Cell Tissue Organ Spatial resolution ( m) LV Wang, S Hu, Science 335, 458, 202; LV Wang, Nature Photon 3, 503, 2009 Outline Motivations and challenges Photoacoustic tomography Photoacoustic computed tomography Photoacoustic microscopy Time-reversal wavefront engineering Compressed ultrafast photography Page 0

11 33 OILAB.SEAS.WUSTL.EDU 32 OILAB.SEAS.WUSTL.EDU 3 OILAB.SEAS.WUSTL.EDU Phase conjugate mirror Time-Reversed Ultrasound-Encoded (TRUE) Optical Focusing Recorded hologram Time-reversed optical focus [X Xu, H Liu], LV Wang, Nature Photon 5, 54, 20; [Y Liu, P Lai], C Ma, X Xu, AA Grabar, LV Wang, Nature Comm 6, 5904, 205; Featured by Nature, 58, 58, 205 Speckle Pattern Concentration by Photoacoustic Wavefront Shaping (PAWS) Initial speckle pattern Single-spot focus Linear x nonlinear intensity enhancement = 60 x 00 = 6000X Acoustic diffraction Optical diffraction [PX Lai, LD Wang, JW Tay], LV Wang, Nature Photon 9, 26, 205 LD Wang, C Zhang, LV Wang, Phys Rev Lett 3, 7430, 204 Outline Motivations and challenges Photoacoustic tomography Photoacoustic computed tomography Photoacoustic microscopy Time-reversal wavefront engineering Compressed ultrafast photography Page

12 36 OILAB.SEAS.WUSTL.EDU 35 OILAB.SEAS.WUSTL.EDU Refraction Fluorescence 34 OILAB.SEAS.WUSTL.EDU Reflection Racing Watch a Flying Laser Pulse with Single-Shot Compressed Ultrafast Photography at 00 Billion Frames/Second [L Gao, J Liang], C Li, LV Wang, Nature 56, 74, 204 Video slowdown: 0 billion X 0 mm Watch a Flying Superluminal Mach Cone with Single-Shot Compressed Ultrafast Photography at 00 Billion Frames/Second Supersonic Mach cone Superluminal Mach cone [J Liang, C Ma, L Zhu], LV Wang, unpublished Single-Shot Compressed Ultrafast Photography: 00 Billion Frames per Second Streak camera shearing S(t) Object with light intensity I(x, y, t) CCD integration T Camera lens Digital micromirror device coding C(x, y) Wide-open entrance slit Objective Tube lens Beam splitter Energy matrix: E(x, y) = TSCI(x, y, t) [L Gao, J Liang], C Li, LV Wang, Nature 56, 74, 204; Comments by B Pogue, Nature 56, 46 Page 2

13 39 OILAB.SEAS.WUSTL.EDU 38 OILAB.SEAS.WUSTL.EDU 37 OILAB.SEAS.WUSTL.EDU Normalized intensity Normalized intensity Boost from 00 Billion to 0 Trillion Frames per Second 00 GHz 2.3 THz 0 mm 0 mm 0 [L Gao, J Liang], C Li, LV Wang, Nature 56, 74, 204 J Liang,..., LV Wang, unpublished Simulated Ultrafast Imaging of Action Potential Propagation: Fastest 2-KHz Commercial Camera versus the Proposed MHz CUP Camera Liren Zhu, LV Wang, unpublished FINANCIAL INTEREST Microphotoacoustics, Inc. Financial Interest Disclosure and Funding Sources ACTIVE GRANTS NIH DP EB06986: NIH Director s Pioneer Award Program Directors: Richard Conroy/Ravi Basavappa NIH R0 CA86567: NIH Director s Transformative Research Award Program Directors: Bob Nordstrom/Ravi Basavappa NIH R0 EB06963: Ring PACT Program Director: Richard Conroy NIH U0 NS090579: BRAIN Initiative Program Director: Ned Talley March of Dimes: Prematurity Birth Program Director: Joe Leigh Simpson Page 3

14 Noise equivalent molar concentration (M) 42 OILAB.SEAS.WUSTL.EDU NEC of hemoglboin (M) NEN of hemoglboin (mol/voxel) 4 OILAB.SEAS.WUSTL.EDU 40 OILAB.SEAS.WUSTL.EDU Further Information YouTube videos on Photoacoustic tomography Web at Books Photoacoustics@gmail.com Relocation to Caltech HIRING: Postdocs Students Technicians Noise Equivalent Concentration or Number 0-3 Microbubble 0-4 MB 0-5 Hb EB 0-6 IRDye Melanin ICG 0-8 mcherry RFP Optical diffusion limit y x 6.3 AR-PAM PAMac EGFP irfp GNB GNC SWNT GNR Molar extinction coefficient (cm - /M) 0-6 SW-PAM JJ Yao, LV Wang, Photoacoustics doi:0.06/j.pacs OR-PAM Imaging depth (mm) Page 4

15 45 OILAB.SEAS.WUSTL.EDU 44 OILAB.SEAS.WUSTL.EDU 43 OILAB.SEAS.WUSTL.EDU Absorption coefficient (cm - ) Endogenous Absorbers Melanin DNA HbO HbR 2 RNA Lipid 0 - Bilirubin MbO 2 MbR Water Wavelength (nm) JJ Yao, LV Wang, Photoacoustics doi:0.06/j.pacs Scalability of Resolution and Penetration Acoustic spatial resolution Acoustic bandwidth Acoustic penetration limit Acoustic bandwidth Penetration limit = Constant 200 Spatial resolution Photoacoustic Conversion Efficiency and SNR Photoacoustic wave from a sphere Thermal expansion coefficient Compressibility 8 mbars mk Noise equivalent pressure ~ sub mbar SNR at photoacoustic source ~ 0 5 Attenuation over a 0 2 voxel range ~ 0 3 SNR at tissue surface ~ 0 2 Page 5

16 48 OILAB.SEAS.WUSTL.EDU mm Normalized PA amplitude 47 OILAB.SEAS.WUSTL.EDU 46 OILAB.SEAS.WUSTL.EDU Fractional change in s ig n a l a m p litu d e (% ) Photoacoustic and Fluorescence Detection of Calcium-Sensitive Protein GCaMP5G in the Fruit Fly Brain In Vivo Odor on P h o to a c o u s tic F lu o re s c e n c e T im e (s e c.) RY Zhang, B Rao, HY Rong, B Raman@WUSTL, LV Wang, unpublished Photoacoustic Microscopy of Calcium-Sensitive Protein GCaMP5G in the Fruit Fly Brain In Vivo RY Zhang, B Rao, HY Rong, B Raman@WUSTL, LV Wang, unpublished Photoacoustic Computed Tomography of the Whole Brain of a Rat In Vivo mm Li Lin, Lei Li, Liren Zhu, Cheng Ma, Konstantin Maslov, LV Wang, unpublished 0 Page 6

17 OILAB.SEAS.WUSTL.EDU Correlation coefficient 49 OILAB.SEAS.WUSTL.EDU Photoacoustic Imaging of Hemoglobin and Glucose Metabolism in Mouse Brain In Vivo with Electric Forepaw Stimulation Analogous to fmri 570 nm (Isosbestic wavelength) Analogous to PET 478 nm (Glucose analog) Right paw stimulation Left paw stimulation Right paw stimulation Left paw stimulation mm 0 0% Photoacoustic change Glucose analog: 2-NBDG = 2-deoxy-2-[N-(7-nitrobenz-2-oxa-,3-diazol-4-yl)amino]-D-glucopyranose [J Yao, J Xia], K Maslov, M Naziriavanaki, V Tsytsarev, AV Demchenko, LV Wang, NeuroImage 64, 257, 203; Collaboration: AV UMSL Noninvasive In Vivo Photoacoustic Tomography of Resting-State Functional Connectivity in Mouse Brain Paxinos atlas Photoacoustic image [M Nasiriavanaki, J Xia], H Wan, A Bauer, J Culver@WUSTL, LV Wang, PNAS, 2, 204 Page 7

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